The mitochondrial ATP synthase is responsible for the synthesis of more than 90% of cellular ATP in the eukaryotic cell under aerobic conditions. ATP is the energy currency that the cell uses for nearly all energy requiring processes such as muscle contraction (as in the heart) and biosynthetic reactions. This has been a major impetus for the intense studies on this enzyme. But recent studies have revealed even greater cause to study the ATP synthase. The ATP synthase has been implicated in a number of other critical processes that are either related or unrelated to the activity of synthesis of ATP. The ATP synthase is a potential target to cure cancer either by eliciting apoptosis or by modulating angiogenesis, to treat Lupus, to extend the life- time of humans either by increasing the metabolic rate thereby providing caloric restriction or by modulating known receptors involved in extended life-span, to prevent heart disease, to treat eye disease, such as glaucoma or macular degeneration, to treat obesity, and to treat bacterial infections such as those caused by Mycobacterium tuberculosis or the opportunistic bacterium, Pseudomonas aeruginosa. The immediate goal of this project is to understand the structure/function relationship of the ATP synthase and to identify critical structural regions of the ATP synthase, which if modulated, inactivate or impair the enzyme. The long-term goal is to target these regions for rational drug design to identify new antibiotics or drugs. There are two objectives for this project. The first objective is to identify and understand molecular structural features critical for the coupling of the flow of protons with the synthesis of ATP by the mitochondrial ATP synthase. The second objective is to identify potential targets in the ATP synthase for rational based drug design. These objectives will be obtained with the same set of experiments. Specifically, regions will be identified within, or associated with, the gamma-subunit, which have strict structural requirements and if perturbed, reduces the efficiency of the coupling of the ATP synthase. This analysis will be done using structure-based mutagenesis studies followed by biochemical and biophysical analysis of the mutant proteins and by structure determination of the ATP synthase. Future studies will target these regions for the development of antibiotics, which either perturb the coupling efficiency or block rotation of the gamma-subunit of the ATP synthase from pathogenic bacteria.